There are currently more than 18 Helicobacter species described and the numbers are increasing. Many of these bacteria are not culturable by standard laboratory methods, since they require microaerophilic conditions and also the presence of hydrogen gas for growth. Many laboratories classify gram-negative, urease-positive, spiral bacteria isolated from gastric specimens as H. pylori without further work-up. Consequently, the incidence and pathogenicity of Helicobacter species other than H. pylori is relatively unclear. Non-culturable Helicobacter-like organisms have been reported in human gastric biopsies, and there are reports of patients with a positive urease breath test with negative gastric biopsy cultures for H. pylori. It is not known whether or not these cases are due to a Helicobacter species other than H. pylori. Additionally, we isolated a strain of Flexispira rappini, a species that is closely related to Helicobacter, in blood. Most Helicobacter species are closely related by 165 RRNA sequencing and possess a rapid urease activity similar to H. pylori. The urease enzyme is strongly expressed and has been considered as a target for vaccine development against H. pylori. However, the database of Helicobacter species urease DNA sequences is limited to three species other than H. pylori. Using polymerase chain reaction (PCR) with degenerate primers, we were able to amplify a region of the ureB gene from H. hepaticas and F. rappini. These PCR products were cloned and sequenced. Analysis of these sequences revealed considerable variability when compared with available sequences. The ureB gene should prove to be a useful PCR amplification target for identification and differentiation between fastidious or non- culturable Helicobacter species. Furthermore, this information should be useful in studies of the prevalence and clinical significance of Helicobacter species other than H. pylori.